Application Note

Single-pass tangential flow filtration

Introduction
Tangential flow filtration (TFF) is widely used in the biopharmaceutical industry for downstream processing
applications. Typical TFF steps concentrate product through volume reduction, and buffer exchange through
diafiltration, to achieve high yields. TFF typically utilizes ultrafiltration membranes ranging from 1-1000 kD
NMWL to retain different size molecules. Traditional TFF requires multiple passes through a system using a pump
to drive feed through a filter and sending the retentate back to a tank for another pass through the system.

Single-pass TFF runs at constant operating conditions throughout the process, simplifies the required hardware,
allows higher concentration factors and higher product recovery without significant dilution by reducing hold-up
volume, and reduces the risk of product damage associated with recirculating TFF operations. Single-pass TFF is also
a continuous process and can be run together with another step. This is convenient to reduce volumes to eliminate
tank bottlenecks and reduce column sizes, especially in existing facilities where space may be limited.

Principles of single-pass TFF

Traditional TFF operates in batch mode, where the feed/
retentate is recirculated through the filter assembly
(Figure 1A). Typically, TFF cassettes operate in parallel,
with multiple passes through membranes required to
achieve the desired concentration.
EMD Millipore single-pass TFF is a different application
of an existing technology (Figure 1B). Single-pass TFF
uses existing Pellicon 2 or Pellicon 3 cassettes
with standard holders. The TFF step is sufficiently
concentrated after a single pass through the filter
assembly such that retentate recycle is not required.
The basic single-pass TFF underlying principal is that
increased residence time in the feed channel results in
increased conversion. Increased residence time can be
accomplished by reducing flow rate or increasing path
length in a serial configuration. Configuring TFF cassettes
in series can improve conversion. Cassettes in series have
a higher mass transfer when compared to parallel
configurations at equivalent residence times
(Figures 1C, 1D).

Retentate
Feed Figure 1.
Compared to traditional
Feed Feed TFF configuration (A),
Permeate a single-pass TFF
Tank Retentate Tank
Retentate configuration (B) does
1C Serial Configuration not recycle the retentate.
Conversion increases with
residence time with a
Permeate Permeate
serial feed channel
Feed Feed Feed Retentate configuration (C) offering
Cassette Cassette
Pump Pump a slight advantage over a
parallel configuration (D),
due to higher flow
1A Batch 1B Single-Pass velocities and mass transfer.
1D Parallel Configuration Permeate not shown.

EMD Millipore is a division of Merck KGaA, Darmstadt, Germany

 Applications of single-pass TFF
Single-pass TFF has several applications, including:
Product concentration/volume reduction: single-pass In-line dilutions/de-salting: Single-pass TFF can be
TFF can be used in-between other unit operations to used for in-line desalting before ion exchange chro
reduce intermediate pool volumes. In turn, this can matography, membrane adsorbers, or virus prefilters
debottleneck a process limited by tank volumes and/ without expanding the pool volume by dilution
or reduce column sizes and/or number of cycles (Figure 2B).
required for downstream chromatography
Final formulation/concentration (post-batch UF/DF):
(Figure 2A).
single-pass TFF reduces working volume limitations
compared to traditional TFF, allowing the process to
achieve higher final concentrations and minimizing
post-use recovery dilution (Figure 2C).

Buffer
Figure 2. Buffer STEP 1
Retentate Retentate
Applications for single-pass
TFF set up for product Feed
Retentate
concentration and volume Feed
reduction (A), inline dilutions Feed
and de-salting (B), and final Permeate
concentration (C).
Permeate Permeate

STEP 2

2A Volume reduction 2B De-Salting 2C Final Concentration

Evaluation of single-pass TFF at small scale

Bench scale experiments using single-pass TFF are
typically performed with three identical Pellicon mini
cassettes in conventional holder(s) (Figure 3A). Specially
designed single-pass TFF diverter plates are used to
allow the three cassettes to be configured in series
within a single holder (Figure 3B and Figure 3C). This
experimental set-up allows the operator to evaluate the
performance of one, two and three-section single-pass
TFF processes at the same time. Alternatively, three
Pellicon Mini holders can be configured in series.

Figure 3B. P1 P2 P3
Bench-scale set up for
single-pass TFF system us- Feed
Tank
ing diverter plates
Retentate
Valve
Pf Section Section Section
0.1 m2 0.1 m2 0.1 m2

Feed
Pump

Separate permeate lines for each section

Figure 3C.
A diverter plate for bench-
scale single-pass TFF set up Figure 3A.
Bench-scale set-up for single-pass TFF system

2
 n
Q
Trials typically require 1-2 L of feed material, and
4-6 hours of run time. The experiment is run in either i
total recycle mode or as a true single pass, depending i=1
Yn =
on the volume of feed available. If the experiment is run QF
in total recycle mode, a well-mixed tank is required to
ensure proper mixing of the concentrated retentate with Where Yn is the conversion for an n-section SPTFF
recycled permeate. process, Q i is the permeate flow measured for sections 1
through i, and Q F is the total feed flow into the system
Single-pass TFF trials require measurement of feed and measured as inlet or sum of retentate and permeate.
retentate pressures and retentate and permeate flow The volume reduction factor (VRF) is defined as:
rates, at varying feed flow rates. Typically, the trial starts VRF= 1/(1-Y)
at feed flux rates of 1 L/min/m2, and is stepped down to By conservation of mass, the steady-state retentate
near 0.1 L/min/m2, until the desired target conversion concentration is expressed as:
is reached or a gel point is reached and the product
Cret = CFeedVRF R
cannot be further concentrated. Each feed flow
point is stabilized for 5-10 minutes prior to taking Evaluation of single-pass TFF with model feed of 20g/L
measurements. Longer times are required at lower and 75g/L demonstrates how the conversion increases
flow rates to allow for displacement of channel volumes. with decreasing feed flow rate, and increases with the
number of sections in series (Figure 4A). Measured
It is convenient to plot the experimentally obtained retentate concentrations confirm predicted
conversion Y=QPtotal/QF vs the feed flux JF. For any concentrations using the mass balance. (Figure 4B).
number of sections in series, one uses the total permeate
in calculating Y and the total area in calculating JF.

100%
90% 18-20 g/L 1-section
18-20 g/L 2-section Figure 4A.
80% 18-20 g/L 3-section
75 g/L 1-section
Example of feed flux
70%
75 g/L 2-section excursion
60%
Conversion

75 g/L 3-section

50%
40%

30%
20%
10%
0%
0.0 0.5 1.0 1.5 2.0 2.5 3.0

Feed Lpm/m2 total

250

Section 3 Predicted Figure 4B.

200
Comparison of measured
Section 3 Measured
Concentration (g/L)

vs predicted retentate
150 concentrations

100

50

0.0 0.5 1.0 1.5

Feed Flow Rate (L/min/m2)

Scaling up single-pass TFF

14 120%

12
100%
Figure 5.
Scale-up uses existing holders and cassettes, including Cleaning of single-pass
10
Pellicon membranes that are commercially available 80% TFF series
NWP (lmh/psi@25C)

and requires no additional customized equipment. 8

NWP/NWPpre

60%

Cleaning of single-pass TFF systems can be performed 6

40%
in single-pass mode without a recirculation loop using 4

typical cleaning agents (Figure 5). Cleaning does not 2

20%

require different pump or piping.

0 0%
Pre Post-IgG Clean

NWP section 1 NWP section 2 NWP section 3 NWP/NWPpre average

3
To scale up single-pass TFF, first define the feed 100%

18-20 gL 1-section
flow (equals batch volume divided by the desired 90%
18-20 g/L 2-section Figure 6.
process time) and target conversion or concentration. 80% 18-20 g/L 3-section Using the feed flux
75 g/L 1-section
excursion graph
Figure 6 shows an example in which greater than 52% 70% 75 g/L 2-section
to determine the
75 g/L 3-section
conversion was required, with a feed flow of 25 Lpm. 60% required feed flux for

Conversion
1 section (light blue)
Using the data in Figure 6, a 52% conversion (red line) 50% Target Conversion
and 2 section
40%
gives feed flux values of 0.6 Lpm/m2 for 1 section, 0.7 (dark blue).
30%
Lpm/m2 for 2 section, and 0.8 Lpm/m2 for 3 section
20%
single-pass TFF.
10%

Next, define the required membrane area by dividing 0%

0.0 0.5 1.0 1.5 2.0 2.5 3.0

the desired feed flow rate by the feed flux. In the Example of required feed flux for 2 section system
Feed Lpm/m2 total

Figure 5 example, the calculation for a 2 section process

is: 25 L pm 0.7 Lpm/m2 = 36m2 total area. This
corresponds to a two section process with 18m2
area per section.

Case study at Summary

manufacturing scale
An example of a single-pass TFF application for
an existing manufacturing facility where improved
cell culture titers placed increased demand on
downstream operations and tank capacities was
reported by Teske et. al. (Biot. Prog. 26(4):1068-72).
In this process, the product pool following virus
filtration exceeded the available tank capacity by
14%. The use of 4m2 of cassettes in a single section met
the 20 L/min flow and volume reduction requirements.
Use of Pellicon cassettes in a single-pass
tangential flow filtration mode is an effective
way of concentrating biomolecules. Small-scale
evaluations are simple to execute and analyze. The
power of implementing single-pass TFF lies in its ability
to reduce in-process volumes in a simple and easy-
to-use step. Single-pass TFF enables higher final
formulations and allows facilities to meet the demands
of higher titer processes without major investments in
new equipment.

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Lit No. AN5572EN00 Rev. A DP-SBU-12-07385 12/2014
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